Deciphering how neural circuits within the mammalian brain give rise to perception, cognition, and behavior is central to understanding how the nervous system functions. Neural circuits operate over a vast range of spatial and computational scales, from high-level circuits that integrate information across multiple brain regions, to microcircuits that perform simple input/output transformations within a specialized brain structure. Each level of analysis is important for formulating responses to environmental conditions. However, studying a specific neural circuit is extremely difficult, as most nervous system structures contain many types of neurons with inextricably intertwined axons and dendrites. To overcome this obstacle, the glycoprotein (G)-deleted rabies vector system was developed to identify direct synaptic inputs to a particular neuronal population. By pseudo typing the G-deleted rabies vector with a foreign envelope protein, such as EnvA, the vector selectively transduces target neurons genetically engineered to express the EnvA receptor. If these cells also express rabies glycoprotein the vector travels retrograde exactly one synaptic step and transduces direct presynaptic neurons. The rabies genome can be altered to encode any gene of interest, including fluorescent proteins to reveal the cytoarchitecture of presynaptic cells, or neuroscience tools (e.g., calcium indicators or light-gated ion channels) to monitor or manipulate circuit activity. Thus, the G-deleted rabies vector system allows the fine- scale manipulation of specific cell types within a circuit, allowing investigators to test hypotheses linking these circuts to behavior. This technology has revolutionized the study of neuronal circuits, creating high demand for these cutting-edge reagents. Laboratories that focus on understanding neural circuits, however, typically do not have the resources or expertise to produce high quality rabies vectors or associated helper vectors that are necessary to perform these experiments. Because of this, the Salk Institute's Gene Transfer, Targeting, and Therapeutics (GT3) Core, which currently generates the rabies vectors, is inundated with requests for ready-to- inject viral reagents and demand exceeds production capacity. This R24 application proposes to expand the GT3 Core's capacity for maintaining, propagating, and distributing all G-deleted rabies vector variants and helper vectors (Aim 1). The GT3 Core will also incorporate newly developed tools into the technology platform as they are innovated (Aim 2). Establishing this central rabies production facility will lower reagent costs (through economies of scale) and improve the reproducibility of study findings. Between-lab cost sharing mechanisms will enable the distribution of small aliquots, facilitating pilot experiments and removing the greatest barrier to technology uptake by new laboratories. Newly generated reagents will be immediately distributed to the neuroscience community without publication restrictions, thereby speeding the pace of discovery. These efforts will broaden the impact of this technology and ensure that neuroscientists studying circuits are equipped with the most modern analytic tools.